Cleavable solid phases for isolating nucleic acids

Abstract

Solid phase materials for binding nucleic acids and methods of their use are disclosed. The materials feature a cleavable linker portion which can be cleaved to release bound nucleic acids. The solid phase materials comprise a solid support portion comprising a matrix selected from silica, glass, insoluble synthetic polymers, and insoluble polysaccharides to which is attached a nucleic acid binding portion for attracting and binding nucleic acids, the nucleic acid binding portion (NAB) being linked by a cleavable linker portion to the solid support portion. Preferred nucleic acid binding portions comprise a ternary or quaternary onium group. The materials can be in the form of microparticles, fibers, beads, membranes, test tubes or microwells and can further comprise a magnetic core portion. Methods of binding nucleic acids using the cleavable solid supports are disclosed as are their use in methods of isolating or purifying nucleic acids.

1 . A solid phase for binding nucleic acids comprising: 
 a solid support portion comprising a matrix selected from silica, glass, insoluble synthetic polymers, and insoluble polysaccharides to which is attached on a surface;    a cleavable linker portion to the solid support portion, and    a nucleic acid binding portion for attracting and binding nucleic acids linked to the cleavable linker portion.    
     
     
         2 . The solid phase of  claim 1  wherein the nucleic acid binding portion is selected from a ternary sulfonium group of the formula SR 2   +  X −  where R is selected from C 1 -C 20  alkyl, aralkyl and aryl groups, a quaternary ammonium group of the formula NR 3   +  X −  wherein R is selected from C 4 -C 20  alkyl, aralkyl and aryl groups, and a quaternary phosphonium group PR 3   +  X −  wherein R is selected from C 1 -C 20  alkyl, aralkyl and aryl groups, and wherein X is an anion.  
     
     
         3 . The solid phase of  claim 2  wherein the nucleic acid binding portion is a quaternary ammonium group and the R groups each contain from 4-20 carbon atoms.  
     
     
         4 . The solid phase of  claim 2  wherein the nucleic acid binding portion is a quaternary phosphonium group and the R groups each contain from 1-20 carbon atoms.  
     
     
         5 . The solid phase of  claim 4  wherein each R group is a butyl group.  
     
     
         6 . The solid phase of  claim 1  wherein the solid support portion comprises an insoluble synthetic polymer.  
     
     
         7 . The solid phase of  claim 1  wherein the solid support portion comprises a glass matrix.  
     
     
         8 . The solid phase of  claim 1  wherein the solid support portion comprises a silica matrix.  
     
     
         9 . The solid phase of  claim 1  wherein the cleavable linker portion further comprises one or more connecting portions.  
     
     
         10 . The solid phase of  claim 1  further comprising a magnetically responsive portion.  
     
     
         11 . The solid phase of  claim 1  wherein the cleavable linker portion is cleaved hydrolytically.  
     
     
         12 . The solid phase of  claim 11  wherein the hydrolytically cleavable linker portion is an ester or thioester group.  
     
     
         13 . The solid phase of  claim 1  wherein the cleavable linker portion is cleaved reductively.  
     
     
         14 . The solid phase of  claim 1  wherein the cleavable linker portion comprises a triggerable dioxetane ring.  
     
     
         15 . The solid phase of  claim 1  wherein the cleavable linker portion comprises an electron rich alkene which is cleaved by conversion to a thermally unstable dioxetane.  
     
     
         16 . The solid phase of  claim 1  wherein the cleavable linker portion is cleaved enzymatically.  
     
     
         17 . The solid phase of  claim 16  wherein the cleavable linker portion comprises an acridan ketene dithioacetal which is cleaved by reaction with a peroxidase and a peroxide.  
     
     
         18 . The solid phase of  claim 16  wherein the cleavable linker portion comprises an ester which is cleaved by a hydrolase enzyme or an esterase enzyme.  
     
     
         19 . The solid phase of  claim 16  wherein the cleavable linker portion comprises an amide which is cleaved by a protease enzyme.  
     
     
         20 . The solid phase of  claim 16  wherein the cleavable linker portion comprises a peptide which is cleaved by a peptidase enzyme.  
     
     
         21 . The solid phase of  claim 16  wherein the cleavable linker portion comprises a glycoside which is cleaved by a glycosidase enzyme.  
     
     
         22 . The solid phase of  claim 12  wherein the cleavable linker portion comprises a thioester having the formula:  
       
         
           
           
               
               
           
         
       
       wherein Q is P or N and R is alkyl of 1-20 carbons.  
     
     
         23 . The solid phase of  claim 22  wherein the cleavable linker portion comprises a thioester having the formula:  
       
         
           
           
               
               
           
         
       
     
     
         24 . The solid phase of  claim 1  wherein the cleavable linker portion is an alkylene group of at least one carbon atom bonded to a trialkylphosphonium or triarylphosphonium nucleic acid binding portion and is cleavable by means of a Wittig reaction with a ketone or aldehyde.  
     
     
         25 . The solid phase of  claim 24  wherein the cleavable linker portion has the formula  
       
         
           
           
               
               
           
         
       
     
     
         26 . The solid phase of  claim 2  wherein the nucleic acid binding portion of the solid phase is a ternary sulfonium group of the formula SR 2   +  X −  where R is selected from C 1 -C 20  alkyl, aralkyl and aryl groups, and wherein X is an anion.